There is a fundamental gap in our understanding of which cell types contribute to residual viremia in HAART treated people. Our laboratory's long-range goal is to understand HIV-1 persistence in patients that are optimally treated with highly active anti-retroviral therapy (HAART). We expect that a greater understanding of HIV persistence will inform the design of therapeutic strategies that will result in long-term remissio or cure. As the next step towards this goal, the objective of this application is to identify celluar sources of residual viremia in optimally treated people and to characterize the residual virus. Our central hypothesis is that residual virus is generated in part by latently infected hematopoietic progenitor cells. The rationale of the proposed work is that identifying cellular reservoirs and viral sub-types that contribute to persistence will yield mechanistic insights into the establishment of latency and accelerate the discovery of disease-modifying therapies. We plan to test our central hypothesis and accomplish the objective of this application by pursuing the following four specific aims: (1) Establish the extent to which bone marrow progenitors harbor HIV genomes in optimally treated people on HAART. We hypothesize, based on preliminary studies, that HSPCs are an inducible reservoir of HIV that can spread infection to T cells. We will test this by measuring the amount of reactivatable HIV proviral DNA in HSPCs from a cohort of optimally treated donors on HAART, including donors who have been on HAART since primary HIV infection. (2) Identify which chemokine receptor is primarily utilized by viruses to enter HSPCs. Our working hypothesis, based on preliminary data, is that HSPCs from HAART-treated donors will harbor dual or CXCR4-tropic HIVs. To test this, we will characterize full-length viral envelopes from these cells and use functional assays to assign tropism. (3) Characterize the contribution of infected HSPCs to residual viremia in optimally treated people on HAART. Our preliminary data led us to hypothesize that reactivated virus from infected HSPCs will seed the periphery. To test this, we will isolate and sequence residual virus and determine based on sequence comparisons to what extent HSPCs versus other possible reservoirs can be identified as the cellular source of this virus. (4) We will determine the mechanisms by which quiescent viral DNA in HSPCs can be reactivated to eliminate latently infected cells. Viral latency in HSPCs has not been well characterized. We will examine the cellular and viral factors that contribute to latency in HSPCs to aid in the selection or development of drugs that can purge latently infected HSPCs. This contribution will be significant because identifying the relevant cellular reservoirs and defining the mechanisms of latency is an important initial step toward the development of better treatment strategies for HIV infected people. If our hypothesis is proven correct, this work will refocus efforts to identify therapeutic targets for eradication of cellular reservoirs to long-lived hematopoietic progenitor cells. PUBLIC HEALTH RELEVANCE: The proposed research is relevant to public health because HIV is an incurable, pandemic virus that has infected millions of people globally and that continues to infect nearly 40,000 people each year in the United States. The outcomes of the proposed research are expected to have an important positive impact by the identification of cellular reservoirs from which latent HIV-1 can cause resurgent viremia and by the identification of potential therapeutic targets for new drug development.